Transition-state structure for the hydronium-ion-promoted hydrolysis of α-d-glucopyranosyl fluoride

Abstract

The transition state for the hydronium-ion-promoted hydrolysis of α-d-glucopyranosyl fluoride in water has been characterized by combining multiple kinetic isotope effect measurements with theoretical modelling. The measured kinetic isotope effects for the C1-deuterium, C2-deuterium, C5-deuterium, anomeric carbon-13, and ring oxygen-18 are 1.219 ± 0.021, 1.099 ± 0.024, 0.976 ± 0.014, 1.014 ± 0.005, and 0.991 ± 0.013, respectively. The transition state for the hydronium ion reaction is late with respect to both C–F bond cleavage and proton transfer.

C2-Oxyanion Neighboring Group Participation: Transition State Structure for the Hydroxide-Promoted Hydrolysis of 4-Nitrophenyl α-d-Mannopyranoside

“…Anomeric carbon KIEs ( k 12 / k 13 ) for the reactions of glycopyranosides in solution are typically in the range of 1.005 to 1.030. Reactions occurring via dissociative transition states exhibit anomeric 13 C-KIE values that are closer to unity; − for instance, the reported 13 C-KIEs for spontaneous hydrolysis of α- d -glucopyranosyl 4-bromo­isoquinolinium bromide and the acid-catalyzed reaction of the anomeric methyl d -xylopyranosides are 1.005 ± 0.002 and 1.006 ± 0.001, respectively. In contrast, the anomeric 13 C-KIE for the base-catalyzed hydrolysis of PNPMan is 1.026 ± 0.006 (Table ), which is in the range typically associated with S N 2 reactions on glycosides that proceed via ‘exploded’ transition states .…”

“…Substitution reactions at the anomeric center of glycosides and other glycosyl derivatives occur through a spectrum of mechanisms. , Owing to the ability of the lone pair on oxygen to stabilize developing charge, reactions that tread the borderline of S N 1 ( A N * D N ) ,,− and S N 2 ( A N D N ) ,,, processes are most common, with the existence of a discrete pyranosylium ion intermediate contingent upon the degree of nucleophile and solvent participation. An S N i (‘internal return’) mechanism ( D N * D h * A N ) has been reported for cases involving an intimate complex of the substrate’s leaving group and a preassociated nucleophile. , Reactions proceeding through such processes have been studied extensively in solution, and closely related counterparts have been identified in various enzymic processes, catalyzed by glycosidases, glycosyltransferases, and carbohydrate phosphorylases (e.g., S N 1, , S N 2, and S N i).…”

Both Chemical and Non-Chemical Steps Limit the Catalytic Efficiency of Family 4 Glycoside Hydrolases

“…We noted no observable activity with Zn 2+ , but Sr 2+ (at a concentration of 1 mM) displays a relative activity of 50% compared to that of Mn 2+ (at the same concentration, data not shown). We initially planned to use α- d -galactopyranosyl fluoride and the appropriate isotopologues as substrates for the measurement of relative rate constants by 19 F NMR spectroscopy: ,, a choice that was based on literature reports that glycosyl fluorides are universally good substrates for the corresponding glycosidase. For example, Williams and Withers stated in their review articles that “... there are no known examples of glycosidases that cannot process the glycosyl fluoride that corresponds to the substrate” .…”

“…The Mn 2+ activated GH4 α-galactosidase from C. freundii efficiently hydrolyzes aryl α-Dgalactopyranoside with no effect of leaving group ability on either the first-order (k cat ) or the second-order (k cat /K m ) enzymatic rate constants. 7 In order to utilize a 19 F NMR methodology 15,19,20 for the measurement of competitive rate constants, we decided to use a diamagnetic divalent metal cation to activate the enzyme rather than paramagnetic Mn 2+ . Therefore, we tested the relative activity of MelA, in comparison to manganous ion, in the presence of Zn 2+ and Sr 2+ using UV−vis spectroscopy with 4-nitrophenyl α-Dgalactopyranoside as the substrate.…”

Glycoside hydrolase stabilization of transition state charge: new directions for inhibitor design

“…24,38,45,46 Therefore, we decided to use 19 F NMR spectroscopy to measure isotopologue ratios (R, the ratio of heavy-to-light isotopologues for the remaining starting material) as the reaction progressed (F, the fraction of reaction for the light isotopologue) to evaluate the secondary deuterium and 13 C-KIEs. 47,48 We note that because our syntheses involved enantioselective catalysis our covalent inhibitors are not enantiopure, as also would be the case with chiral pool starting materials, such as galactose. We corrected our R and F values, assuming that the enzyme would have no reaction with the small quantity of L-carbasugar in the reaction mixture (details in the Experimental section and Tables S1 and S2 in ESI †).…”